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Calcium specificity signaling mechanisms in abscisic acid signal transduction in Arabidopsis guard cells.

Brandt B, Munemasa S, Wang C, Nguyen D, Yong T, Yang PG, Poretsky E, Belknap TF, Waadt R, Alemán F, Schroeder JI - Elife (2015)

Bottom Line: In intact guard cells, abscisic acid (ABA) enhances (primes) the Ca(2+)-sensitivity of downstream signaling events that result in activation of S-type anion channels during stomatal closure, providing a specificity mechanism in Ca(2+)-signaling.However, the underlying genetic and biochemical mechanisms remain unknown.Moreover, we demonstrate an unexpected interdependence of the Ca(2+)-dependent and Ca(2+)-independent ABA-signaling branches and the in planta requirement of simultaneous phosphorylation at two key phosphorylation sites in SLAC1.

View Article: PubMed Central - PubMed

Affiliation: Division of Biological Sciences, Cell and Developmental Biology Section, University of California, San Diego, San Diego, United States.

ABSTRACT
A central question is how specificity in cellular responses to the eukaryotic second messenger Ca(2+) is achieved. Plant guard cells, that form stomatal pores for gas exchange, provide a powerful system for in depth investigation of Ca(2+)-signaling specificity in plants. In intact guard cells, abscisic acid (ABA) enhances (primes) the Ca(2+)-sensitivity of downstream signaling events that result in activation of S-type anion channels during stomatal closure, providing a specificity mechanism in Ca(2+)-signaling. However, the underlying genetic and biochemical mechanisms remain unknown. Here we show impairment of ABA signal transduction in stomata of calcium-dependent protein kinase quadruple mutant plants. Interestingly, protein phosphatase 2Cs prevent non-specific Ca(2+)-signaling. Moreover, we demonstrate an unexpected interdependence of the Ca(2+)-dependent and Ca(2+)-independent ABA-signaling branches and the in planta requirement of simultaneous phosphorylation at two key phosphorylation sites in SLAC1. We identify novel mechanisms ensuring specificity and robustness within stomatal Ca(2+)-signaling on a cellular, genetic, and biochemical level.

No MeSH data available.


Related in: MedlinePlus

Analysis of ABA activation of S-type anion currents in PP2C quadruple mutant guard cells at low [Ca2+]cyt.(A and B) ABA application in WT and abi1-2/abi2-2/hab1-1/pp2ca-1 guard cells with [Ca2+]cyt buffered to a resting level of 0.1 μM does not result in large S-type anion current activation. Typical current traces (A), average steady-state currents in response to applied voltages (±SEM), and numbers of individual measured guard cells are shown (B). Several error bars are not visible, as these were smaller than the illustrated symbols.DOI:http://dx.doi.org/10.7554/eLife.03599.006
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fig2s1: Analysis of ABA activation of S-type anion currents in PP2C quadruple mutant guard cells at low [Ca2+]cyt.(A and B) ABA application in WT and abi1-2/abi2-2/hab1-1/pp2ca-1 guard cells with [Ca2+]cyt buffered to a resting level of 0.1 μM does not result in large S-type anion current activation. Typical current traces (A), average steady-state currents in response to applied voltages (±SEM), and numbers of individual measured guard cells are shown (B). Several error bars are not visible, as these were smaller than the illustrated symbols.DOI:http://dx.doi.org/10.7554/eLife.03599.006

Mentions: Members of the clade A of the PP2C class play important roles as negative regulators of ABA signaling (Cutler et al., 2010) and were shown to inhibit CPK-activation of SLAC1 in oocytes (Geiger et al., 2010; Brandt et al., 2012). To determine whether these PP2Cs function in the ABA-triggered enhancement of the [Ca2+]cyt-sensitivity in guard cells, we performed whole-cell patch-clamp analysis using a plant line carrying T-DNA insertion mutations in the key ABA signaling PP2Cs ABI1, ABI2, HAB1, and PP2CA (abi1-2/abi2-2/hab1-1/pp2ca-1). Surprisingly, in abi1-2/abi2-2/hab1-1/pp2ca-1 quadruple mutant guard cells, strong Ca2+-activated S-type anion currents were observed even without pre-exposure to ABA (Figure 2A–D). At low 0.1 μM [Ca2+]cyt S-type anion channels did not show significant activation in the pp2c quadruple mutant compared to WT (Figure 2—figure supplement 1A,B; p = 0.294 at −145 mV). These findings provide genetic evidence for first genes that are essential for the ABA-triggered Ca2+ sensitivity priming in guard cells and show that these PP2Cs provide a mechanism ensuring specificity in Ca2+ signal transduction.10.7554/eLife.03599.005Figure 2.In protein phosphatase 2C (PP2C) quadruple mutant plants, Ca2+ activation of S-type anion currents is constitutively primed.


Calcium specificity signaling mechanisms in abscisic acid signal transduction in Arabidopsis guard cells.

Brandt B, Munemasa S, Wang C, Nguyen D, Yong T, Yang PG, Poretsky E, Belknap TF, Waadt R, Alemán F, Schroeder JI - Elife (2015)

Analysis of ABA activation of S-type anion currents in PP2C quadruple mutant guard cells at low [Ca2+]cyt.(A and B) ABA application in WT and abi1-2/abi2-2/hab1-1/pp2ca-1 guard cells with [Ca2+]cyt buffered to a resting level of 0.1 μM does not result in large S-type anion current activation. Typical current traces (A), average steady-state currents in response to applied voltages (±SEM), and numbers of individual measured guard cells are shown (B). Several error bars are not visible, as these were smaller than the illustrated symbols.DOI:http://dx.doi.org/10.7554/eLife.03599.006
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4507714&req=5

fig2s1: Analysis of ABA activation of S-type anion currents in PP2C quadruple mutant guard cells at low [Ca2+]cyt.(A and B) ABA application in WT and abi1-2/abi2-2/hab1-1/pp2ca-1 guard cells with [Ca2+]cyt buffered to a resting level of 0.1 μM does not result in large S-type anion current activation. Typical current traces (A), average steady-state currents in response to applied voltages (±SEM), and numbers of individual measured guard cells are shown (B). Several error bars are not visible, as these were smaller than the illustrated symbols.DOI:http://dx.doi.org/10.7554/eLife.03599.006
Mentions: Members of the clade A of the PP2C class play important roles as negative regulators of ABA signaling (Cutler et al., 2010) and were shown to inhibit CPK-activation of SLAC1 in oocytes (Geiger et al., 2010; Brandt et al., 2012). To determine whether these PP2Cs function in the ABA-triggered enhancement of the [Ca2+]cyt-sensitivity in guard cells, we performed whole-cell patch-clamp analysis using a plant line carrying T-DNA insertion mutations in the key ABA signaling PP2Cs ABI1, ABI2, HAB1, and PP2CA (abi1-2/abi2-2/hab1-1/pp2ca-1). Surprisingly, in abi1-2/abi2-2/hab1-1/pp2ca-1 quadruple mutant guard cells, strong Ca2+-activated S-type anion currents were observed even without pre-exposure to ABA (Figure 2A–D). At low 0.1 μM [Ca2+]cyt S-type anion channels did not show significant activation in the pp2c quadruple mutant compared to WT (Figure 2—figure supplement 1A,B; p = 0.294 at −145 mV). These findings provide genetic evidence for first genes that are essential for the ABA-triggered Ca2+ sensitivity priming in guard cells and show that these PP2Cs provide a mechanism ensuring specificity in Ca2+ signal transduction.10.7554/eLife.03599.005Figure 2.In protein phosphatase 2C (PP2C) quadruple mutant plants, Ca2+ activation of S-type anion currents is constitutively primed.

Bottom Line: In intact guard cells, abscisic acid (ABA) enhances (primes) the Ca(2+)-sensitivity of downstream signaling events that result in activation of S-type anion channels during stomatal closure, providing a specificity mechanism in Ca(2+)-signaling.However, the underlying genetic and biochemical mechanisms remain unknown.Moreover, we demonstrate an unexpected interdependence of the Ca(2+)-dependent and Ca(2+)-independent ABA-signaling branches and the in planta requirement of simultaneous phosphorylation at two key phosphorylation sites in SLAC1.

View Article: PubMed Central - PubMed

Affiliation: Division of Biological Sciences, Cell and Developmental Biology Section, University of California, San Diego, San Diego, United States.

ABSTRACT
A central question is how specificity in cellular responses to the eukaryotic second messenger Ca(2+) is achieved. Plant guard cells, that form stomatal pores for gas exchange, provide a powerful system for in depth investigation of Ca(2+)-signaling specificity in plants. In intact guard cells, abscisic acid (ABA) enhances (primes) the Ca(2+)-sensitivity of downstream signaling events that result in activation of S-type anion channels during stomatal closure, providing a specificity mechanism in Ca(2+)-signaling. However, the underlying genetic and biochemical mechanisms remain unknown. Here we show impairment of ABA signal transduction in stomata of calcium-dependent protein kinase quadruple mutant plants. Interestingly, protein phosphatase 2Cs prevent non-specific Ca(2+)-signaling. Moreover, we demonstrate an unexpected interdependence of the Ca(2+)-dependent and Ca(2+)-independent ABA-signaling branches and the in planta requirement of simultaneous phosphorylation at two key phosphorylation sites in SLAC1. We identify novel mechanisms ensuring specificity and robustness within stomatal Ca(2+)-signaling on a cellular, genetic, and biochemical level.

No MeSH data available.


Related in: MedlinePlus